Evidence has accumulated indicating that a highly conserved hydrophobic domain usually near the amino terminus of viral transmembrane proteins plays critical roles in the infection process. These viral fusion sequences insert into and destabilize the cell membrane and act in concert with the viral and cellular proteins and lipids to form a pore through which the viral nucleic acid can pass. Site-directed mutagenesis has shown that replacement of key residues in the influenza virus hemagglutinin (HA2) and the HIV glycoprotein 41 (gp41) fusion sequences can dramatically affect viral fusion. Synthetic "fusion peptides" (FP) corresponding to the fusion sequences, induce lipid mixing and lysis with liposomes and cell membranes. Some studies with synthetic FP representing the mutated viral fusion sequences have better defined the structural origins of the altered viral fusion. However there is still limited information concerning the relationship of viral FP structure to their mechanism of action. This research proposal is based on the hypothesis that understanding of the role of the fusion sequence in virus-cell fusion can be enhanced by defining the detailed structural and functional changes brought about by replacement of key residues in synthetic fusion peptides. The objective is to determine the structural characteristics of viral FP that are necessary for fusion competence. Using FP based on mutated viral sequences, we will seek correlations between their altered activity and membrane bound structure. The influenza, HIV and variant FP will be tested for lytic and fusogenic activity with red blood cells and liposomes. The nature and extent of the FP and variant perturbation of membrane lipid will be investigated with 31P-NMR, polarized Fourier Transform Infrared (FTIR) and Electron Spin Resonance (ESR) spectroscopy. The conformation, orientation and topography of fusion peptides in membranes or membrane mimicking solvents will also be examined with Circular Dichroism (CD), FTIR, and ESR spectroscopy. FTIR of 13C-labeled peptides will be used to assess local conformations. Based on these structural studies, we will then construct molecular models for the interaction of each FP and variant with membrane lipids.